Skin Regenerative Potential of Cupuaçu Seed Extract (Theobroma grandiflorum), a Native Fruit from the Amazon: Development of a Topical Formulation Based on Chitosan-Coated Nanocapsules.

Scarless skin regeneration is a challenge in regenerative medicine. Herein, we explore the regenerative potential of a Cupuaçu seed extract (Theobroma grandiflorum) to develop an innovative skin regeneration formulation based on chitosan-coated nanocapsules. Cupuaçu seed extract significantly stimulated cell proliferation and migration. A reparative gene expression profile could be verified following extract treatment, which included high levels of MKI67, a cellular proliferation marker, and extracellular matrix genes, such as ELN and HAS2, which code for elastin and hyaluronic acid synthase 2. Formulations with Cupuaçu seed extract successfully entrapped into nanocapsules (EE% > 94%) were developed. Uncoated or coated nanocapsules with low-molecular-weight chitosan presented unimodal size distribution with hydrodynamic diameters of 278.3 ± 5.0 nm (PDI = 0.18 ± 0.02) and 337.2 ± 2.1 nm (PDI = 0.27 ± 0.01), respectively. Both nanosystems were physically stable for at least 120 days and showed to be non-irritating to reconstructed human epidermis. Chitosan coating promoted active penetration into undamaged skin areas, which were still covered by the stratum corneum. In conclusion, the present study demonstrated for the first time the biotechnological potential of the frequently discarded Cupuaçu seed as a valuable pharmaceutical ingredient to be used in regenerative skin products.

Desenvolvimento de achocolatados em pó com adição de subprodutos de frutas, processados por spray dryer e com modificador reológico / Development of powdered chocolate added with fruit byproducts, processed by spray dryer and rheological modifier

O achocolatado em pó é um dos derivados do cacau com maior inserção econômica e cultural em diversos países. O objetivo deste estudo foi avaliar a adição de ingredientes diferenciados nesse tipo de produto, como modificadores reológicos e fruta, bem como à alteração no tipo de cacau utilizado, ocasionando mudanças sensoriais e nutricionais positivas ao produto. O fruto sugerido neste estudo foi o cupuaçu (Theoboroma grandiflorum), fruto típico da região Norte, que apresenta excelente qualidade nutricional. Foram desenvolvidas 7 formulações de achocolatado por método convencional após simples mistura (padrão, com cacau alcalino, com cacau orgânico, com polpa de cupuaçu, com amido pré-gelatinizado, com amido pré-gelatinizado + polpa de cupuaçu, com goma guar, com goma guar + polpa de cupuaçu) e 4 formulações processadas por spray dryer após a simples mistura (padrão, com polpa de cupuaçu, com amido pré-gelatinizado, com amido + polpa de cupuaçu). Todas as formulações foram avaliadas quanto à composição nutricional, calorimetria exploratória de varredura (DSC), análises físico-químicas, reológica, quantificação dos compostos fenólicos e avaliação da capacidade antioxidante por métodos in vitro. Em seguida foi realizada análise sensorial com as formulações: padrão, com polpa de cupuaçu e com amido + polpa de cupuaçu. O achocolatado padrão apresentou tempo de mistura de 38 min, o que foi utilizado como parâmetro para as demais formulações. Os achocolatados que continham polpa de cupuaçu apresentaram maior teor proteico (14,5 a 16,3 g/100g) quando comparados com o padrão (13,6 g/100g). Todos os achocolatados apresentaram umidade entre 1,2% e 3,7%, e atividade de água entre 0,13 e 0,57, considerados microbiologicamente estáveis, sendo bom para a vida útil do produto. Os achocolatados obtiveram tempo de molhabilidade entre 07:15 min e 15:06 min; solubilidade de 1,56 IR% a 7,44 IR%; tamanho de partícula variando entre 0,216 mm e 0,347 mm (partículas finas). O uso do spray dryer não teve impacto significativo nas características físicas das formulações, assim como a utilização dos diferentes tipos de cacau não afetou a composição nutricional e qualidade física dos achocolatados. Houve aumento (p< 0,05) para o tempo de molhabilidade e solubilidade do achocolatado com cacau orgânico em comparação com o padrão (13:30 e 9:33 min; 2,64 e 1,56 IR%, respectivamente). A transição vítrea variou entre 35,2 a 35,7 mW enquanto o ponto de carbonização ficou entre 237,4 a 243,6 mW, indicando que a adição dos agentes espessantes e/ou do cupuaçu não interferiu (p<0,05) na análise térmica dos achocolatados. Todos os achocolatados diluídos em leite apresentaram-se como pseudoplásticos, com aumento de viscosidade nas menores temperaturas, conforme esperado. O achocolatado com cacau orgânico apresentou o maior teor de compostos fenólicos (8,27 mg AG g-1) enquanto observou-se redução no conteúdo de fenólicos nos produtos processados por spray dryer. Os achocolatados apresentaram capacidade antioxidante entre 31,76 µMETrolox/g e 75,62 µMETrolox/g, pelo método do DPPH. A adição do cupuaçu levou ao aumento da capacidade de sequestro de radicais DPPH quando comprados com o padrão (p<0,05). Não foi observada diferença significativa pelo método FRAP. A avaliação sensorial obteve aceitação situada na região positiva da escala (5 a 7). Os achocolatados formulados apresentam formulações adequadas a sua comercialização, com agregação de valor nutricional e econômico

The chocolate powdered is a cocoa-derived with greater economic and cultural integration in several countries. The objective of this study was to evaluate the addition of different ingredients in this type of product, such as rheological modifiers and fruit, as well as the change in the type of cocoa used, causing positive sensory and nutritional changes to the product. The fruit suggested in this study was cupuassu (Theoboroma grandiflorum), a typical fruit from the northern region, which has excellent nutritional quality. Seven powdered chocolate formulations were developed by conventional method after simple mixing (standard, with alkaline cocoa, with organic cocoa, with cupuassu pulp, pre-gelatinized starch, pre-gelatinized starch + cupuassu pulp, guar gum, with guar gum + cupuassu pulp) and 4 formulations processed by spray dryer after simple mixing (standard, with cupuassu pulp, pre-gelatinized starch, starch + cupuassu pulp). All formulations were evaluated for nutritional composition, differential scanning calorimetry (DSC), physicochemical, rheological analyzes, quantification of phenolic compounds and antioxidant capacity evaluation by in vitro methods. Then, sensory analysis was performed with the formulations: standard, with cupuassu pulp and starch + cupuassu pulp. The standard powdered chocolate had a mixing time of 38 min, which was used as parameter for the other formulations. The powdered chocolate containing cupuassu pulp had higher protein content (14.5 to 16.3 g / 100g) when compared to the standard (13.6 g / 100g). All powdered chocolate presented humidity between 1.2% and 3.7%, and water activity between 0.13 and 0.57, considered microbiologically stable, wich is good for the shelf life of the product. The powdered chocolate obtained wettability time between 07:15 min and 15:06 min; solubility from 1.56 IR% to 7.44 IR%; particle size ranging from 0.216 mm to 0.347 mm (fine particles). The use of the spray dryer had no significant impact on the physical characteristics of the formulations, as well as the use of different types of cocoa did not affect the nutritional composition and physical quality of the powdered chocolate. There was an increase (p <0.05) for the time of wettability and solubility in chocolate powdered formulated with organic cocoa when compared to the standard (9:33 and 13:30 min; IR 2.64 and 1.56%, respectively). The glass transition ranged from 35.2 to 35.7 mW while the carbonization point ranged from 237.4 to 243.6 mW, indicating that the addition of thickening agents and / or cupuassu did not interfere (p <0.05) in the thermal analysis of powdered chocolate. All powdered chocolate when diluted in milk presented as pseudoplastics, with viscosity increase at lower temperatures, as expected. Chocolate powdered with organic cocoa presented the highest content of phenolic compounds (8.27 mg AG g-1) whereas there was a reduction in phenolic content in products processed by spray dryer. The powdered chocolates presented antioxidant capacity between 31.76 µMETrolox / g and 75.62 µMETrolox / g, by the DPPH method. The addition of cupuassu led to increased ability to sequester DPPH radicals when compared to the standard (p <0.05). No significant difference was observed by the FRAP method. Sensory evaluation was accepted in the positive region of the scale (5 to 7). The formulated powdered chocolates have appropriate formulations for marketing, with added nutritional and economic value

Monitoring and Optimization of Cupuaçu Seed Fermentation, Drying and Storage Processes.

Cupuaçu [Theobroma grandiflorum (Wild ex Spreng.) K. Schum] seeds have been employed for a long time in the Amazon region for food purposes. Similar to cocoa, processed cupuaçu pulp and seeds can be used to produce juices, ice creams, confectionary products and cupulate®, which is a similar product to chocolate. However, its market penetration requires the mastery of all processing stages to improve the food quality and safety and to make possible an efficient technology transfer to the local small farmers and communities. Based on the above, the current research work aimed at monitoring and optimizing the consecutive fermentation and drying processes of cupuaçu seeds over 7 days each, as well as storage for 90 days. A greenhouse structure incorporating the fermenter and solar drying terrace was designed to be inexpensive, versatile, easily scalable, and easy to maintain and operate by the local small farmers after a short period of training. This research effort also aimed at giving a vision for the future creation of an integrative and sustainable cupuaçu system covering the economic, social, cultural and environmental vectors. The experimental design comprised 5 batches of 100 kg of seeds each. Several microbiological and physicochemical parameters were performed and correlated with processing variables. Microbiological parameters encompassed viable counts of mesophilic microorganisms, coliforms, yeasts, and molds, whereas physicochemical measures included fermentation and drying temperature, pH, acidity, dry matter, ashes, water activity, color, total proteins, lipids and carbohydrates, and energy. The average seed fermentation temperature varied from ca. 28 to 44 °C, reaching the maximum on day 3 and a final value of ca. 31 °C. Regarding solar drying, the average seed temperatures ranged from ca. 24 °C (at the end) to 39 °C on day 3, and an initial value of ca. 29 °C. The average final seed pH value of drying was 5.34 and was kept during storage. During storage, results demonstrated the existence of significant correlations among several experimental parameters under scrutiny. Finally, bean viable counts obtained during storage unfolded acceptable values of total mesophilic bacteria well below the maximum limit. Viable counts of yeast and molds were generally found between 3 and 4 log(CFU/gsample), and total coliforms were also detected, although both were at acceptable levels and well beneath the established maximum limits for food safety.

Preparing multicomponent snack bars based on tapioca flour, Brazil nut, and regional fruits / Preparación de barras multicomponentes a base de harina de tapioca, castaña de brasil y frutos regionales

ABSTRACT This study aimed to develop and assess the physicochemical, sensory parameters, and shelf life estimation of multicomponent snack bars based on tapioca flour, Brazil nut, and açaí or cupuassu pulp. The physicochemical composition of açaí- and cupuassu-flavored snack bars had, respectively, 0.92 and 0.99% ash, 19.22 and 17.02% lipids, 3.02 and 3.03% protein, 1.06 and 1.69% fiber, and 448 and 436 kcal/100 g energy value. The shear stress test showed the consumer needs to bite more strongly to break the açaí-flavored bar. The opposite was observed in the hardness test, in which the bite compression force during mastication was greater for the cupuassu-flavored bar. The bars had water activity below 0.6, which denotes microbiological stability. The sensory analysis ranked the bars between "liked slightly" and "liked very much," which was confirmed by the acceptability index above 75% for all attributes assessed. According to the results a significant increase in water activity over storage was observed suggest the packaging used in the tests did not present a satisfactory barrier to water vapor permeability. Only water activity was used to estimate shelf life, which was determined as 58 days and 49 days for the açaí- and cupuassu-flavored bars, respectively. Thus, the snack bars represent an alternative for athletes as well as individuals with celiac disease since they are gluten free.

RESUMEN El objetivo de este trabajo fue desarrollar y evaluar los parámetros físicos, físico-químicos, microbiológicos, sensoriales y la vida útil en estante de barras a base de harina de tapioca, castaña de Brasil y pulpa de açaí o cupuaçu. En cuanto a la composición físico-química, las barras multicomponentes sabor açaí y cupuaçu presentaron, respectivamente, 0,92 y 0,99% de cenizas, 19,22 y 17,02% de lípidos, 3,02 y 3,03% proteínas, 1,06 y 1,69% de fibras y 448 y 436 kcal/100g de valor energético. La prueba de cizallamiento y dureza mostraron que el consumidor necesita una fuerza de mordida mayor para romper la barra sabor açaí. El comportamiento contrario fue observado en la prueba de dureza donde la fuerza de compresión de la mordida, durante la masticación, fue mayor en la barra sabor cupuaçu. Para el análisis sensorial se observó que las barras evaluadas recibieron notas situadas entre las categorías "me gustó ligeramente" y "me gustó mucho", resultado comprobado por el índice de aceptabilidad con valores superiores al 75% para todos los atributos evaluados. De acuerdo con los resultados, se observó un aumento significativo en la actividad de agua durante el almacenamiento, lo que sugiere que el embalaje utilizado en las pruebas no presentó una barrera satisfactoria para la permeabilidad al vapor de agua. Para la estimación de vida de estante sólo la actividad de agua fue utilizada para los cálculos, siendo el tiempo de vida de estante determinado en 58 días para la barra sabor açaí y 49 días para la barra sabor cupuaçu. Así, las barras multicomponentes elaboradas representam una alternativa para atletas, así como para portadores de la enfermedad celíaca, visto la ausencia de gluten en su composición.

Digestibilidade, comportamento ingestivo e desempenho de ovinos alimentados com resíduos de agroindústrias processadoras de frutas / Digestibility, intake behavior, and performance of sheep fed residues from fruit processing agro-industries

Objetivou-se avaliar o consumo, a digestibilidade e o desempenho de ovinos, arraçoados com dietas isoproteicas, contendo 75% de resíduos da agroindústria frutífera (abacaxi, acerola, maracujá e cupuaçu), uma dieta com feno de tifton e 25% de concentrado, em delineamento inteiramente ao acaso. A participação do resíduo de maracujá na dieta proporcionou os maiores níveis de consumo de matéria seca (1170,6g d-1) em relação aos resíduos de abacaxi (693,8g d-1), acerola (644,2g d-1), cupuaçu (452,9g d-1) e feno de tifton (962,7g d-1), o que se relaciona aos seus níveis de digestibilidade (77,8; 63,1; 59,1; 61,7 e 71,8%, respectivamente). Tal resultado gerou diferenças significativas, destacando-se o resíduo de maracujá quanto à conversão alimentar (8,3), e o ganho médio diário de peso (141,5g d-1). Apesar de ter tido a mesma conversão observada para o feno de tifton (13,2), apresentou maior ganho médio diário (75,6g d-1). Os demais resíduos apresentaram efeitos sobrepostos, contudo o resíduo de acerola ficou com a pior conversão (51,4) e o menor ganho médio diário (15,6g d-1), sendo esse sem diferença em relação ao cupuaçu (23,0g d-1). Os resíduos podem ser utilizados como alternativa para a alimentação de ovinos.(AU)

The objective was to evaluate the consumption, digestibility and performance of the sheep with iso protein diets, with 75% of residues from fruit agro-industries (pineapple, acerola, passion fruit, and cupuassu) and a diet with tifton hay, and 25% of concentrate, in a totally randomized design. The participation of passion fruit residue on the diet provided the highest consumption levels of dry matter (1170.6g d -1 ) compared with the residues of pineapple (693.8g d -1 ), acerola (644.2g d -1 ), cupuassu (452.9g d -1 ) and tifton hay (962.7g d -1 ), which is related to their levels of digestibility (77.85, 63.14, 59.07, 61.68 and 71.83%, respectively). Such a result generated significant differences with emphasis on passion fruit residue regarding food inversion (8.30) and average daily weight gain (141.50g d -1 ), in spite of having the same conversion detected for tifton hay (13.2), it showed a higher average daily gain (75.6g d -1 ). The other residues showed overlapping effects, however, acerola residue showed the worst conversion level (51.4) and the lowest average daily gain (15.6g d -1 ), and this one showed no difference compared with cupuassu (23.0g d -1 ). The residues can be used as an alternative for sheep feeding.(AU)

Modificações estruturais e reológicas em chocolate amargo devido à alteração do tipo de gordura utilizada / Structural and rheological changes in bitter chocolate due to the change in the type of fat used

O chocolate é conhecido mundialmente, proveniente do fruto do cacaueiro (Theobroma cacao) normalmente consumido em forma de barra, mas também podendo ser usado de inúmeras formas como coberturas, recheios dentre outras. Um importante componente na produção do chocolate é a gordura utilizada, uma vez que esta é responsável pela textura, brilho e características organolépticas do produto. O objetivo deste trabalho foi utilizar a manteiga de cupuaçu (proveniente do fruto da Theobroma grandiflorum) na elaboração de chocolate amargo. Para tanto a manteiga de cacau foi substituída de forma parcial e total. Foram desenvolvidas duas formulações de chocolate padrão com liquor de cacau (P1) e com cacau em pó (P2), e quatro formulações com substituição parcial da manteiga de cacau por manteiga de cupuaçu a partir de P2(F1 e F2) e de P1 (F3 e F4). As amostras elaboradas e os ingredientes (líquor de cacau, cacau em pó, manteiga de cacau e manteiga de cupuaçu) foram avaliadas por análise térmica (DSC-Differential Scanning Calorimetry), reologia, tamanho de partícula, composição em ácidos graxos e em triacilgliceróis, índice de temperagem e índice de resfriamento, bem como testes de acompanhamento de 112 dias como cor, atividade de água e textura. A manteiga de cupuaçu apresentou maior quantidade de ácido oleico quando comparada com a manteiga de cacau, aproximadamente 11,5%, e também características reológicas diferentes dos padrões: tensões iniciais variaram de 3,4 ± 0,3 a 7,9 ± 2,0 Pa para as amostras e 2,9 ± 1,4 a 6,2 ± 0,7 Pa para os padrões; viscosidade de 1,6 ± 0,1 a 2,9 ± 0,4 Pa*s para as amostras e 1,9 ± 0,8 a 2,9 ± 0,9 Pa*s para os padrões; tamanho de partícula das amostras de 21 ± 2 a 22 ± 2 µm, padrões de 20 ± 2 a 34 ± 4 µm. Durante os 112 dias de estudo de prateleira: Aw variou de 0,405 ± 0,03 a 0,424 ± 0,02 nas amostras e 0,399 ± 0,03 no padrão; textura variou de 16,3 ± 1,2 a 31,6 ± 2,0 N para as amostras e 25,9 ± 3,0 a 28,6 ± 7,2 N para os padrões; WI variou de 24,1 ± 0,6 a 25,4 ± 0,3 para as amostras e 23,0 ± 0,4 a 23,9 ± 0,8 para os padrões; ΔE variou de 0,4 a 2,2 para as amostras e de 0,5 a 1,2 para os padrões. Concluiu-se que: A variação do líquor de cacau para cacau em pó acarretou aumentando do tempo total de processo em aproximadamente 15 min. Os chocolates com maior teor de manteiga de cupuaçu apresentaram aumento em triacilglicerol C54, com redução de POP e POS. A faixa de fusão dos chocolates com maiores porcentagens de manteiga de cupuaçu (F3 e F4) foi maior do que para chocolates formulados apenas com manteiga de cacau (P1 e P2). A manteiga de cupuaçu tem relação direta com a queda da tensão inicial e da viscosidade (sem diferença significativa, p<0,05) nos chocolates produzidos. O baixo ponto de fusão do ácido oleico contido na manteiga de cupuaçu alterou a temperatura final e o valor do índice de temperagem nas amostras com maior teor de manteiga de cupuaçu (F3 e F4)

The chocolate is known worldwide, coming from the fruit of the cacao (Theobroma cacao) normally consumed in the form of a bar, but also can be used in countless ways like coverings, fillings among others. An important component in the production of chocolate is the fat used, since it is responsible for the texture, brightness and organoleptic characteristics of the product. The objective of this work was to use cupuassu butter (from the fruit of Theobroma grandiflorum) in elaboration of bitter chocolate. For this purpose, the cocoa butter was partially and totally replaced. Two formulations of standard chocolate with cocoa liquor (P1) and with cocoa powder (P2) were developed, and four formulations with partial replacement of cocoa butter by cupuassu butter from P2 (F1 and F2) and P1 (F3 and F4). The elaborated samples and the ingredients (cocoa liquor, cocoa powder, cocoa butter and cupuassu butter) were evaluated by DSC, rheology, particle size, fatty acid composition and triacylglycerol, temperature index and cooling index, as well as follow-up tests of 112 days such as color, water activity and texture. The cupuassu butter presented a higher amount of oleic acid when compared to cocoa butter, approximately 11.5%, and also different rheological characteristics of the standards: initial tensions ranged from 3.4 ± 0.3 to 7.9 ± 2, 0 Pa for the samples and 2.9 ± 1.4 to 6.2 ± 0.7 Pa for the standards; viscosity of 1.6 ± 0.1 to 2.9 ± 0.4 Pa*s for the samples and 1.9 ± 0.8 to 2.9 ± 0.9 Pa*s for the standards; sample particle size from 21 ± 2 to 22 ± 2 µm, patterns from 20 ± 2 to 34 ± 4 µm. During the 112 days of shelf study: Aw ranged from 0.405 ± 0.03 to 0.424 ± 0.02 in the samples and 0.399 ± 0.03 in the standard; texture ranged from 16.3 ± 1.2 to 31.6 ± 2.0 N for the samples and 25.9 ± 3.0 to 28.6 ± 7.2 N for the standards; WI ranged from 24.1 ± 0.6 to 25.4 ± 0.3 for the samples and 23.0 ± 0.4 to 23.9 ± 0.8 for the standards; ΔE ranged from 0.4 to 2.2 for the samples and from 0.5 to 1.2 for the standards. It was concluded that: The variation of cocoa liquor to cocoa powder increased the total process time by approximately 15 min. The chocolates with higher content of cupuassu butter presented increase in triacylglycerol C54, with reduction of POP and POS. The melting range of chocolates with higher percentages of cupuassu butter (F3 and F4) was higher than for chocolates formulated with cocoa butter alone (P1 and P2). Cupuassu butter is directly related to the drop in initial tension and viscosity (without significant difference, p <0.05) in the chocolates produced. The low melting point of the oleic acid contained in the cupuassu butter altered the final temperature and the temperature index value in the samples with the highest cupuassu butter content (F3 and F4)

Selection of Lactic Acid Bacteria with Probiotic Potential Isolated from the Fermentation Process of "Cupuaçu" (Theobroma grandiflorum).

In the present study, nine lactic acid bacteria isolated from the fermentation process of "cupuaçu" (Theobroma grandiflorum) were selected for probiotic use. In vitro (resistance to gastrointestinal environment, in vitro antagonism and co-aggregation with pathogens) and in vivo (intestinal colonization and ex vivo antagonism in germ-free mice, cumulative mortality, translocation to liver and spleen, histopathological examination of liver and ileum and mRNA cytokine gene expression during an experimental infection with S. Typhimurium) assays were used. Among the nine Lactobacillus strains isolated from the "cupuaçu" fermentation, L. plantarum 81 and L. plantarum 90 were selected as potential probiotics based on better results obtained in in vitro evaluations (production of diffusible inhibitory compounds and co-aggregation) as well as in vivo experiments (resistance to gastrointestinal environment, ex vivo antagonism, higher survival after enteropathogen challenge, lower hepatic translocation of enteropathogen, lower histopathological lesions in ileum and liver and anti-inflammatory pattern of immunological response). Concluding, L. plantarum 81 and L. plantarum 90 showed in vitro and in vivo capacities for probiotic use through different mechanisms of protection and its origin would allow an easier adaptation in an alimentary matrix for its administration.

Hidrolisado proteico da semente de cupuaçu como fonte de peptídeos inibidores da enzima conversora da angiotensina I / Hydrolyzed cupuassu seed protein as a source of angiotensin I- converting enzyme inhibitory peptides

Peptídeos com ação inibitória da enzima conversora da angiotensina I (ECA) podem ser obtidos a partir de diversos alimentos e exercer efeito anti-hipertensivo. O cupuaçu (Theobroma grandiflorum S.), fruto nativo da Amazônia, possui sementes comestíveis contendo proteína de reserva similar à do cacau (Theobroma cacao L.), as quais parecem ser fonte de peptídeos inibidores da ECA. Desse modo, o objetivo deste trabalho foi investigar in vitro a ocorrência de peptídeos inibidores da ECA no hidrolisado proteico da semente de cupuaçu obtido por ação da Alcalase. O hidrolisado revelou o desaparecimento de proteínas entre 27 a 180 kDa, incluindo as globulinas, e o surgimento daquelas abaixo de 15 kDa após 2 h de hidrólise, indicando a formação de peptídeos menores. O ensaio de atividade utilizando o substrato Abz-FRK(Dnp)-P-OH revelou que o hidrolisado total promoveu 65% de inibição da ECA e esse pool peptídico foi fracionado em cinco frações (F1-F5) por cromatografia em fase reversa (RP-HPLC). Após a etapa de purificação, o monitoramento da inibição apontou, ao final, duas frações (3.2.8 e 3.4.10) com maior atividade inibitória. Oito peptídeos foram identificados por LC-MS/MS, sendo três deles já conhecidos como inibidores da ECA. Outros cinco novos peptídeos identificados (FLEK, GSGKHVSP, LDNK, MVVDKLF e MEKHS) foram sintetizados e tiveram sua ação inibitória validada por ensaios in vitro. O peptídeo GSGKHVSP apresentou a menor IC50 (3,11 µM) e Ki (0,74 µM), sendo um inibidor tipo misto quanto ao seu mecanismo de inibição revelado pelo gráfico de Lineweaver-Burk. Os resultados permitem concluir que o isolado proteico da semente de cupuaçu pode ser uma fonte para obtenção de peptídeos anti-hipertensivos, a despeito de serem necessárias investigações sobre a resistência desses peptídeos à digestão gastrointestinal e a eficácia da inibição in vivo

Peptides with angiotensin I-converting enzyme (ACE) inhibitory activity may be obtained from several foods and cause antihypertensive effect. Cupuassu (Theobroma grandiflorum S.), a native fruit from Amazon, has edible seeds with a storage protein similar to that of cocoa (Theobroma cacao L.) which seems to have incrypted ACE inhibitor peptides. Thus, the aim of this project was to investigate the in vitro formation of ACE inhibitory peptides in protein hydrolysate from cupuassu seeds using Alcalase enzyme. The hydrolysate revealed the disappearance of proteins between 27 and 181 kDa after 2h hydrolysis, including the globulin, and the increase of those below 15 kDa, indicating the formation of peptides. The ACE inhibitory activity assays using the substrate Abz-FRK(Dnp)P-OH revealed the hydrolysate had 65% ACE inhibition and the pool of peptides was fractionated into five fractions (F1-F5) by reversed phase high-performance liquid chromatography (RP-HPLC). After the purification step, two fractions (3.2.8 e 3.4.10) exhibited the highest ACE-inhibitory activity. Eight peptides had been identified by LC-MS/MS and three of them were ACE inhibitors. The other newly identified peptides (FLEK, GSGKHVSP, LDNK, MVVDKLF and MEKHS) were synthesized and in vitro assayed for ACE inhibitory activity. The peptide GSGKHVSP had the lower IC50 (3.11 µM) and Ki (0.74 µM). Lineweaver-Burk plots suggest this peptide is a mixed-type inhibitor according to the inhibition mechanism. The results indicate that protein isolate from cupuassu seeds may be a good protein source of antihypertensive peptides and further investigation is needed in order to evaluate the resistance of these peptides to gastrointestinal digestion and the inhibitory activity in vivo

Leaf anatomical features of three Theobroma species (Malvaceae s.l. ) native to the Brazilian Amazon / Caracteres anatômicos foliares de três espécies de Theobroma (Malvaceae s.l. ) nativas da Amazônia Brasileira

Theobroma species have economic importance due to their use in the cosmetic and food industries, mainly in the production of chocolate. However, the anatomy of their vegetative structures remains poorly studied. The goal of this study was to describe the anatomical features of Theobroma grandiflorum, T. speciosum and T. subincanum to contribute to the biological knowledge of these species, as well as provide support to the biotechnological studies of native fruit plants of the Amazon. Leaves at different developmental stages were collected and analyzed under light microscopy and scanning electron microscopy. Sessile and stalked stellate trichomes and digitiform glandular trichomes were observed in the expanded leaves of T. grandiflorum and T. subincanum. These species were also similar in the morphology of the midrib, the organization of the mesophyll and the presence of starch grains in the midrib pith cells. Claviform glandular trichomes and mucilage cells in the epidermis occurred only in the expanded leaves of T. speciosum. The presence of mucilage secretory trichomes in shoot apices (colleters) of all species is a new finding for the genus Theobroma.

As espécies de Theobroma têm importância econômica devido a sua utilização nas indústrias cosmética e alimentícia, principalmente na produção de chocolate. Entretanto, a anatomia de suas estruturas vegetativas permanece pouco conhecida. O presente estudo teve por objetivo descrever as características anatômicas de Theobroma grandiflorum, T. speciosum e T. subincanum, como contribuição ao conhecimento biológico dessas espécies, bem como, fornecer subsídios aos estudos biotecnológicos de fruteiras nativas da Amazônia. Folhas em diferentes estágios de desenvolvimento foram coletadas e analisadas sob microscopia de luz e eletrônica de varredura. Nas folhas expandidas de T. grandiflorum e T. subincanum foram observados tricomas dos tipos estrelado séssil, pedunculado e glandular digitiforme. Estas espécies também foram similares quanto à morfologia da nervura central, à organização do mesofilo e à presença de grãos de amido no parênquima medular da nervura central. Tricomas glandulares claviformes e células mucilaginosas na epiderme do limbo foliar ocorreram somente em T. speciosum. A presença de tricomas secretores de mucilagem nos ápices vegetativos (coléteres) de todas as espécies estudadas é um novo registro para o gênero Theobroma.

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Chromatin differentiation between Theobroma cacao L. and T. grandiflorum Schum.

A comparative analysis of mitotic chromosomes of Theobroma cacao (cacao) and T. grandiflorum (cupuaçu) was performed aiming to identify cytological differences between the two most important species of this genus. Both species have symmetric karyotypes, with 2n = 20 metacentric chromosomes ranging in size from 2.00 to 1.19 µm (cacao) and from 2.21 to 1.15 µm (cupuaçu). The interphase nuclei of both species were of the arreticulate type, displaying up to 20 chromocentres, which were more regularly shaped in cacao than in cupuaçu. Prophase chromosomes of both species were more condensed in the proximal region, sometimes including the whole short arm. Both species exhibited only one pair of terminal heterochromatic bands, positively stained with chromomycin A (3) , which co-localized with the single 45S rDNA site. Each karyotype displayed a single 5S rDNA site in the proximal region of another chromosome pair. Heterochromatic bands were also observed on the centromeric/pericentromeric regions of all 20 chromosomes of cacao after C-banding followed by Giemsa or DAPI staining, whereas in cupuaçu they were never detected. These data suggest that the chromosomes of both species have been largely conserved and their pericentromeric chromatin is the only citologically differentiated region.

Chromatin differentiation between Theobroma cacao L. and T. grandiflorum Schum

A comparative analysis of mitotic chromosomes of Theobroma cacao (cacao) and T. grandiflorum (cupuaçu) was performed aiming to identify cytological differences between the two most important species of this genus. Both species have symmetric karyotypes, with 2n = 20 metacentric chromosomes ranging in size from 2.00 to 1.19 µm (cacao) and from 2.21 to 1.15 mm (cupuaçu). The interphase nuclei of both species were of the arreticulate type, displaying up to 20 chromocentres, which were more regularly shaped in cacao than in cupuaçu. Prophase chromosomes of both species were more condensed in the proximal region, sometimes including the whole short arm. Both species exhibited only one pair of terminal heterochromatic bands, positively stained with chromomycin A3, which co-localized with the single 45S rDNA site. Each karyotype displayed a single 5S rDNA site in the proximal region of another chromosome pair. Heterochromatic bands were also observed on the centromeric/pericentromeric regions of all 20 chromosomes of cacao after C-banding followed by Giemsa or DAPI staining, whereas in cupuaçu they were never detected. These data suggest that the chromosomes of both species have been largely conserved and their pericentromeric chromatin is the only citologically differentiated region.

Elaboração e caracterização de barras de frutas regionais desidratadas / Preparation and characterization of regional dehydrated fruit bars

O presente trabalho teve por objetivo desenvolver um produto tipo “barra de frutas” utilizando frutas desidratadas na Amazônia. Foram usados frutos popularmente conhecidos como castanha do Brasil, cupuaçu e pupunha de modo a se obter um produto de boa aceitação e qualidade. O produto foi elaborado com frutas desidratadas em estufa de circulação forçada de ar e triturado. Análises físico-químicas, microbiológicas e sensoriais foram realizadas para avaliação da qualidade do produto. Os testes microbiológicos apresentaram um produto dentro dos padrões para consumo. Análises físico-químicas mostraram um produto de elevado teor de proteínas, carboidratos e minerais. Os testes sensoriais mostraram que 96,66% das 50 pessoas entrevistadas aprovaram o produto.

Calogênese do híbrido Theobroma grandiflorum x T. obovatum (Sterculiaceae) / Calogenesis of Theobroma grandiflorum x. T. obovatum hibrid (Sterculiaceae)

O declínio da cacauicultura na amazônia tem sido causado pela doença vassoura-de-bruxa (Crinipellis perniciosa). Um híbrido interespecífico do mesmo gênero (Theobroma grandiflorum x T. obovatum) tem demonstrando resistência a esta doença. Avaliou-se a calogênese deste híbrido com a finalidade de sua propagação massiva. Os explantes usados (tegumento, cotilédone e embrião) foram postos para desenvolver em condições estéreis em meio de cultura LPm mais água de coco (50 ml/l), glucose (20 g/l), 2,4 - D (2 mg/l), Thidiazuron - TDZ nas concentrações de 0, 5 e 10 æg/l, carvão ativado (2 g/l) em gelrite (2 g/l) a um pH de 5,8. Cada concentração de TDZ x tipo de tecido foi considerado um tratamento. As culturas foram mantidas no escuro, à temperatura de 22 ± 2 °C. Os tecidos não mostraram diferenças significativas quanto a proporção do explante recoberto por calos (p= 0,14) embora notou-se que nos cotilédones tenha sido maior. A dosagem de TDZ nitidamente não influenciou no recobrimento de calos (p= 0,94). Quanto a freqüência, os tecidos mostraram diferentes capacidades de desenvolver calos (p= 0,02) sem mostrar-se influenciados pela dosagem de TDZ (p = 0,66). Os explantes de cotilédones foram os que formaram calos em maior freqüência. Houve uma leve tendência de que quanto menor a dosagem de TDZ maior a freqüência de calos obtidos. Não foi constatada interação entre tipo de calo/frasco x dosagem de TDZ (p = 0,32).

Decline of cocoa plantations in Amazonia has been attributed to Witches-broom-disease (Crinipellis perniciosa). An interspecific hybrid (Theobroma grandiflorum x T. obovatum) has demonstrated resistance to this disease. Calogenesis of this hybrid was evaluated, in preparation for its mass propagation. The explants used (tegument, cotyledons and embryos) were developed in sterile conditions in LPm media plus coconut milk (50 ml/l), glucose (20 g/l), 2,4 - D (2 mg/l), Thidiazuron - TDZ at concentrations of 0, 5 e 10 æg/l, and activated charcoal (2 g/l) in gelrite (2 g/l) at pH 5.8. Each concentration of TDZ x kind of explant was considered a treatment and were kept in the dark at a temperature of 22 ± 2 °C. Explants did not show significant differences in the proportion of callus recovery (p= 0,14) although cotyledons did have the highest value. TDZ level did not influence callus recovery (p= 0,94). Different explants resulted in different frequencies of callus development (p= 0,023) but were not been influenced by the doses of TDZ (p = 0,66). Cotyledon explants displayed the highest values for frequency of callus development. There was also a slight trend that lower levels of TDZ resulted in higher frequencies of callus development. There was no interaction between kind of callus/flasks x doses of TDZ (p = 0,32).

Kinetics of flavour and aroma changes in thermally processed cupuaçu (Theobroma grandiflorum) pulp.

Changes in 'fresh' and 'cooked-notes' during thermal treatment of cupuaçu (Theobroma grandiflorum) pulp were evaluated and modelled. Isothermal experiments in the temperature range of 70-98 °C were carried out and a non-linear regression was performed to all data to estimate kinetic parameters. 'Fresh' and 'cooked-notes' change followed simple first-order (Ea = 78-82 kJ·mol-1 , z = 30-31 °C) and reversible first order (Ea = 80-85 kJ·mol-1 ) kinetics, respectively. Although 'cooked-notes' were linearly correlated with 'fresh-notes' (R2 = 0.99), the former was a better indicator for quality degradation. These results are useful to design pasteurisation processes while minimising sensory changes. © 2000 Society of Chemical Industry.

BIOLOGIA DE Conotrachelus humeropictus, FIEDLER, 1940 (COLEOPTERA: CURCULIONIDAE), PRAGA DO CACAUEIRO E DO CUPUAÇUZEIRO NA AMAZÔNIA BRASILEIRA

The biology of Conotrachelus hutneropictus Fiedler, 1940, a serious pest of Theobroma cacao L. and T. grandiflorum (Will, ex Spreng.) Schum, were investigated. The research was conducted in laboratories under controled temperature (27 ± 2oC) and relative humidity (80 ± 10%) conditions, C. hutneropictus fed on young leaves and fruits of cocoa. Its cycle from egg to adult emergency from soil, was estabilished in average from 79 to 151 days, being 108.80 ± 9.44 (X ± SE95) days for males and 156.87 ±16.19 days for females. The pre-oviposition period was 16.20 ± 0.49 days. The oviposition period was 80.50 ± 5.58 days, when each female lays 55 to 153 eggs in average 108.45 ± 6.21 eggs, and the daily oviposition was 1.29 ± 0.03 eggs. For the larval and embrionaire phases inside the cocoa fruits, the insect needed 25 to 39 days, (31.37 ± 10.96 days). After this period, 20.25 ± 1.50 days in the soil were necessary for pre-pupal (6,07 ± 0,06 days) and pupal (9,62 ±0,10 days) development and completphysiologic adult formation (4,56 ±0,11 days). By this time, C. humeropictus becomes more vulnerable to attack of natural enemies, specially fungi Metarhizium anisopliae (Metsch.) Sorok and Beauveria bassiana (Bals.) Vuill. only enemies constated in the researche.

Foram determinados diversos parâmetros da biologia de Conotrachelus hutneropictus Fiedler, importante praga do cacaueiro, Theobroma cacao L. e do cupuaçuzeiro, T. grandiflorum (Will, ex Spreng.) Schum na Amazônia brasileira. A pesquisa foi realizada em laboratório, com fotofase natural de 12 horas, sob condições controladas de temperatura (27 ± 2eC) e umidade relativa (80 ± 10%). C. hutneropictus alimentado com folhas novas e frutos novos de cacueiros, apresentou em média, um ciclo de ovo a emergência do adulto do solo de 79 a 151 dias e em média, 108,80 ± 9,44 dias ( ±95) para machos e 156,87 ±16,19 dias para fêmeas. O período de pré-oviposição foi de 16,20 ± 0,49 dias. O periodo de oviposição foi de 80,50 ± 5,58 dias, colocando cada fêmea, de 55 a 153 ovos, em média 108,45 ± 6,21 ovos, sendo as posturas diárias e constituídas em média por 1,29 ± 0,03 ovos. No interior dos frutos de cacau, o inseto necessitou de 4,73 ± 0,06 dias e 26,64 ±0,17 dias, para o desenvolvimento embrionário e larval, respectivamente. Após esse período, um total de 20,25 ± 1,50 dias se passaram no solo, para o desenvolvimento das fases de pré-pupa (6,07 ± 0,06 dias), pupa (9,62 ±0,10 dias) e completa formação fisiológica do adulto (4,56 ±0,11 dias). Neste interim, C. humeropictus torna-se mais vulnerável ao ataque de seus inimigos naturais, principalmente fungos, dentre os quais Metarhiziutn anisopliae (Metsch.) Sorok. e Beauveria bassiana (Bals.) Vuill., únicos inimigos constatados para a espécie no decorrer da pesquisa.

ESTIMATIVA DA ÁREA FOLIAR E DO PESO DE FOLHAS SECAS DE PLANTAS JOVENS DE CUPUAÇU (Theobroma grandiflorum (WILLD. EX SPRENG.) SCHUM. - STERCULIACEAE) POR MÉTODOS NÃO DESTRUTIVOS

From forty leaves of 2.5 month old cupuassu seedlings, area of the leaf, fresh weight, blade fresh weight, leaf length, blade length, greatest blade width, distance between the base and the greatest width and the dry weight of the leaf, were taken. Using "Stepwise" regression, equations were obtained to estimate the area of the leaf and the dry weight of the leaf. The equation which gave the best fit (log area of the leaf = -0.133 + 1.095 log blade length + 0.872 log blade width, R2 = 0.996) accounted for 92.5% of the data calculated with a deviation of up to 10% of measured areas. The equations studied to estimate the dry weight of the leaf were inefficient. Alternative equations to estimate leaf weight and area of the leaf based on the natural logarithims of easily taken measurements were evaluated.

De 40 folhas de plantas de cupuaçu com 2,5 meses de idade, foram medidos: área de cada folha, peso da folha fresca, peso do limbo fresco, comprimento da folha, comprimento do limbo, maior largura do limbo, distância da base do limbo até a sua maior largura e o peso da folha seca. Entre essas variáveis, com o uso da análise de regressão por passos (stepwise), foram obtidas equações de regressão para estimar a área da folha e o peso da folha seca. Com a equação obtida para a determinação da área da folha (log da área da folha = -0,133 + 1,095 log Comprimento do limbo + 0,872 log Largura do limbo R2= 0,996), conseguiu-se que 92,5% dos dados calculados apresentassem desvios menores do que 10% do valor observado. As equações testadas para a determinação do peso da folha seca mostraram-se ineficientes. Foram avaliadas equações alternativas para o cálculo do peso da matéria seca e da área da folha usando-se o logaritmo natural das medições mais laceis de serem feitas.

A POLINIZAÇÃO MANUAL DO CUPUAÇUZEIRO (Theobroma grandiflorum)

Cupuassu (Theobroma grandiflorum,Sterculiaceae) is an Amazonian tree whose fruits are used to make juice, ice-cream, candies and chocolate. It has gained popularity recently and is considered one of the most profitable crops in the Amazon region. The species is predominantly allogamous and many flowers do not set fruit, even when pollinated with compatible pollen. The rate of natural pollination is low (estimated at 1.6% for pistils pollinated with more than 60 pollen grains). In the present work, some aspects of its pollination were studied. Hand pollination was performed close to anthesis (between 17-18:00 h) and on the morning following anthesis, in the following way: three staminodes of the flower to be pollinated were removed to allow access to the pistil; a stamen was then taken from the donor flower with a pair of tweezers and its anthers were brushed against the stigmatic branches of the receptor flower. Best results with hand pollination were obtained with recently opened flowers and attributed to a high deposition of pollen grains. Controlled pollinations between compatible plants, using a variable number of pollen grains, showed that flowers that received 60 pollen grains had a 20% probability to set fruit; flowers that received more than 400 pollen grains always set fruit. Hand pollination increased the number of harvested fruits. The number of seeds from fruits obtained by natural and hand pollination was similar. Hand pollination, therefore, can be used both for breeding and for yield enhancement of trees that would otherwise have pollination problems.

O cupuaçu (Theobroma grandiflorum,Sterculiaceae) é uma frutífera da Amazônia usada na fabricação de sucos, sorvetes, doces e chocolate. A sua popularidade vem aumentando e atualmente é considerada uma das culturas mais lucrativas da região Amazônica. O cupuaçu é uma espécie predominantemente alógama e muitas flores não chegam a dar frutos mesmo quando polinizadas com póíen compatível. A taxa de polinização natural é baixa (estimado em 1,6% para pístílos polínízados com mais de 60 grãos de pólen). No presente trabalho, foram estudados alguns aspectos da sua polinização. Polinizações manuais foram feitas próximo à antese (entre 17-18:00 h) e na manhã seguinte após a antese, da seguinte maneira: retiraram-se três estaminódios da flor que iria ser polinizada para facilitar o acesso ao pistilo; da flor doadora de pólen retirou-se um estame, cujas anteras foram passadas nos braços estigmáticos. O maior êxito foi obtido nas polinizações feitas próximas à antese, atribuído ao maior número de grãos depositados durante as polinizações. Polinizações controladas entre plantas compatíveis mostraram que as flores que receberam 60 grãos de pólen tiveram 20% de probabilidade de formar fruto; as flores que receberam mais de 400 grãos de pólen, sempre formaram mitos. O uso de polinização manual aumentou o número de frutos maduros colhidos. O número de sementes nos frutos derivados de polinização natural e manual foi similar. A polinização manual poderá ser utilizada tanto no melhoramento como para aumentar a produtividade dos cupuaçuzeiros que estejam com problemas de polinização.